Green and nanoacaricides including essential oil (EO) nanoemulsions are important compounds to provide new, active, safe acaricides and lead to improvement of avoiding the risk of synthetic acaricides. This study was carried out for the first time on eriophyid mites to develop nanoemulsion of garlic essential oil by ultrasonic emulsification and evaluate its acaricidal activity against the two eriophyid olive mites Aceria oleae Nalepa and Tegolophus hassani (Keifer). Acute toxicity of nanoemulsion was also studied on male rats. Garlic EO was analyzed by gas chromatography–mass spectrometry (GC-MS), and the major compounds were diallyl sulfide (8.6%), diallyl disulfide (28.36%), dimethyl tetrasulfide (15.26%), trisulfide,di-2-propenyl (10.41%), and tetrasulfide,di-2-propenyl (9.67%). Garlic oil nanoemulsion with droplet size 93.4 nm was formulated by ultrasonic emulsification for 35 min. Emulsification time and oil and surfactant ratio correlated to the emulsion droplet size and stability. The formulated nanoemulsion showed high acaricidal activity against injurious eriophyid mites with LC₅₀ 298.225 and 309.634 μg/ml, respectively. No signs of nanoemulsion toxicity were noted in treating rats; thus, it may be considered non-toxic to mammals. Stability of garlic oil nanoemulsion, high acaricidal activity, and the absence of organic toxic solvents make the formulation that may be a possible acaricidal product. Results suggest the possibility of developing suitable natural nanoacaricide from garlic oil.